The proposed research plan involves several aspects of (Na plus K ions-ATPase. The first section deals with the partial reactions of the enzyme system, such as Na ion-dependent phosphorylation, Na ion- dependent exchange reaction, and K ion-stimulated dephosphorylation. Preliminary data suggest that there may be two hydrolytic sites on the enzyme and that the transport enzyme lacks substrate specificity not only for the phosphorylation reaction but for exchange activity as well. These studies may help determine a molecular basis for Na ion transport. The second section examines the hypothesis that ethacrynic acid-mediated diuresis involves inhibition of (Na plus K ion)-ATPase. The conflict regarding in vivo effects of ethacrynic acid may be due to inability of previous workers to obtain relatively pure enzyme preparations without deoxycholate and EDTA treatment. Preliminary experiments indicate that in a microsomal preparation from medulla of dog kidney treated with heparin, it is possible to demonstrate inhibition of (Na plus K ion)-ATPase in vivo by a nontoxic dose of ethacrynic acid. Finally, the third section involves (Na plus K ion)-ATPase as a cardiotonic receptor for ouabain. Since the ouabain-enzyme complex dissociates at high K ion concentrations, it is possible that previous workers failed to observe inhibition of (Na plus K ion)-ATPase by ouabain in vivo, as the concentration of K ion was high in their assay system. Therefore, inhibition of some partial reactions of (Na plus K ion)-ATPase in absence of K ion will be determined to assess the effect of ouabain in vivo. Furthermore, the inotropic effect of ouabain may involve inhibition of (Na plus K ion)-ATPase in a specific area of the heart. Hence, (Na plus K ion)-ATPase activity in different areas of the heart will be determined.